Hosono Akira, Mizuguchi Hiroyuki, Hayashi Hideyuki, Goto Masaru, Miyahara Ikuko, Hirotsu Ken, Kagamiyama Hiroyuki
Department of Biochemistry, Osaka Medical College, 2-7 Daigakumachi, Takatsuki 569-8686.
J Biochem. 2003 Dec;134(6):843-51. doi: 10.1093/jb/mvg210.
A subfamily I aminotransferase gene homologue containing an open reading frame encoding 381 amino acid residues (Mr=42,271) has been identified in the process of the genome project of an extremely thermophilic bacterium, Thermus thermophilus HB8. Alignment of the predicted amino acid sequence using FASTA shows that this protein is a member of aminotransferase subfamily Igamma. The protein shows around 40% identity with both T. thermophilus aspartate aminotransferase [EC 2.6.1.1] and mammalian glutamine:phenylpyruvate aminotransferase [EC 2.6.1.64]. The recombinant protein expressed in Escherichia coli is a homodimer with a subunit molecular weight of 42,000, has one pyridoxal 5'-phosphate per subunit, and is highly active toward glutamine, methionine, aromatic amino acids, and corresponding keto acids, but has no preference for alanine and dicarboxylic amino acids. These substrate specificities are similar to those described for mammalian glutamine: phenylpyruvate aminotransferase. This is the first enzyme reported so far that has the glutamine aminotransferase activity in non-eukaryotic cells. As the presence of aromatic amino acid:2-oxoglutarate aminotransferase [EC 2.6.1.57] has not been reported in T. thermophilus, this enzyme is expected to catalyze the last transamination step of phenylalanine and tyrosine biosynthesis. It may also be involved in the methionine regeneration pathway associated with polyamine biosynthesis. The enzyme shows a strikingly high pKa value (9.3) of the coenzyme Schiff base in comparison with other subfamily I aminotransferases. The origin of this unique pKa value and the substrate specificity is discussed based on the previous crystallographic data of T. thermophilus and E. coli aspartate aminotransferases.
在嗜热栖热菌（Thermus thermophilus）HB8的基因组计划研究过程中，已鉴定出一个亚家族I转氨酶基因同源物，其开放阅读框编码381个氨基酸残基（分子量为42,271）。使用FASTA对预测的氨基酸序列进行比对，结果表明该蛋白质是转氨酶亚家族Iγ的成员。该蛋白质与嗜热栖热菌天冬氨酸转氨酶[EC 2.6.1.1]和哺乳动物谷氨酰胺：苯丙酮酸转氨酶[EC 2.6.1.64]的同一性均约为40%。在大肠杆菌中表达的重组蛋白是一种同源二聚体，亚基分子量为42,000，每个亚基含有一个磷酸吡哆醛5'-磷酸，对谷氨酰胺、蛋氨酸、芳香族氨基酸和相应的酮酸具有高活性，但对丙氨酸和二羧酸氨基酸无偏好。这些底物特异性与哺乳动物谷氨酰胺：苯丙酮酸转氨酶的底物特异性相似。这是迄今为止报道的第一个在非真核细胞中具有谷氨酰胺转氨酶活性的酶。由于尚未报道嗜热栖热菌中存在芳香族氨基酸：2-氧代戊二酸转氨酶[EC 2.6.1.57]，因此预计该酶可催化苯丙氨酸和酪氨酸生物合成的最后一步转氨反应。它也可能参与与多胺生物合成相关的蛋氨酸再生途径。与其他亚家族I转氨酶相比，该酶的辅酶席夫碱具有极高（9.3）的pKa值。基于嗜热栖热菌和大肠杆菌天冬氨酸转氨酶先前的晶体学数据，讨论了这一独特pKa值和底物特异性的来源。
